Abstract A HIV-1 Envelope (Env) vaccine that elicits broadly neutralizing antibodies (bnAbs) has remained elusive, in part due to our limited understanding of the mechanisms of bnAb induction during natural infection. Recently developed simian-human chimeric immunodeficiency viruses (SHIVs) bearing Envs from HIV-1 transmitted- founder (TF) viruses associated with bnAb induction in HIV-1 infected adult individuals now provide an experimental model to establish the mechanisms of bnAb development. HIV-1 Env bnAbs develop in ~50% of chronically-infected adults in over 3-5 years of infection, but HIV-1-infected infants and children more frequently develop bnAbs and do so faster than adults. Understanding the cellular and molecular mechanisms of infant bnAb development will facilitate design of vaccine strategies to elicit bnAbs. In a recent study of the single cell transcriptome profile of neonate and adult macaque immune cells after HIV-1 Env vaccination, we found that neonates had a higher frequency of marginal zone (MZ)-like B cells compared to adults. MZ B cells are known to function as innate-like lymphocytes and mount a rapid Ab response to pathogens, but whether neonates predominantly utilize MZ B cells to generate Abs to HIV-1 infection or vaccination is not known. In adults, a key correlate of bnAb induction is the ratio of follicular helper (Tfh) to regulatory (Tfr) CD4 T cell subsets in blood, and Tfh cells are postulated to stimulate bnAb B cells to acquire affinity maturation in germinal centers (GC). Whether Tfh are elevated in blood or lymph nodes of HIV-1-infected infants is unknown. In this grant, we will use blood and immune tissues to interrogate infant immunity to new TF Env SHIVs, and determine the B cell subsets that give rise to NAbs, including bnAbs, and the roles of CD4 T cells in infant NAb induction. The specific aims for this proposal are as follows: Aim 1. Determine the dynamics of virus evolution and immune escape in neonatal compared to adult macaque SHIV infections Aim 2. Define B cell evolution pathways elicited in neonatal SHIV infections Aim 3. Interrogate lymph node GC and blood CD4 T and B cell subsets, and evaluate their function in neonatal SHIV infections